The topography of the active site of Horseradish Peroxidase (HRP) and manganic HRP has been studied with the aid of a spin-labeled analog of benzhydroxamic acid. The electron spin resonance measurements indicated that the nitroxide moiety of this spin label when bound to HRP became highly immobilized. The 2T" varied inversely with temperature, reaching a maximum of 68.25 G at 0 degrees C and a minimum of 46.5 G at 52 degrees C. From the dipolar interaction between iron atom and free radical, the minimum distance between the two spins was estimated to be 14 A. BIBLIOGRAPHIC REFERENCES: Wee, V.T., Feldmann, R.J., Tanis, R.J. and Chignell, C.F.: A comparative study of mammalian erythrocyte carbonic anhydrases employing spin-labeled analogues of inhibitory sulfonamides. Mol. Pharmac. 12: 832-843, 1976.